Molecular topological fractionation of macromolecules

ABSTRACT

A process for characterizing a sample comprising a population of linear macromolecules of interest ( 104 ) and a population of long chain branched macromolecules of interest ( 103 ), the process including four steps. The first step is to provide a flow through separating medium ( 100 ) and a liquid eluant ( 101 ) in which the macromolecules of interest dissolve, the separating medium defining flow through channels ( 102 ), the eluant flow rate and the average diameter of the channels being in a range so that the linear macromolecules of interest elute before the long chain branched macromolecules of interest ( 105 ) ( 106 ). The second step is to introduce a sample into the liquid eluant. The third step is to flow the liquid eluant under pressure through the channels ( 102 ) of the separating medium ( 100 ). The fourth step is to differentiate the linear macromolecules of interest ( 104 ) from the long chain branched macromolecules of interest ( 103 ) based on their successive elution volumes established in the third step such as by determining the refractive index of the successive elution volumes or by subjecting the successive elution volumes to size exclusion chromatography.

TECHNICAL FIELD

The instant invention is in the field of chemical analysis. Morespecifically, the instant invention relates to chemical analysis methodsfor the characterization of branching topology of macromolecules.

BACKGROUND ART

Common polymers, such as polystyrene or polyethylene, theoreticallycomprise extended linear chains of monomers. However, such polymers canalso have branches; see Graessley et al., Macromolecules, Vol. 9, No. 1,1976, p 127. Such branched polymers often have significantly betterprocessing characteristics (especially when the number of monomer unitsin the branch chain exceeds 100 to 300 units) than their linear or shortchain branched counterparts. For example, the melt strength of a longchain branched polymer can be significantly higher than the meltstrength of its linear or short chain branched counterpart of the samemolecular weight and often shear thin to a greater extent, see Macosko,Rheology—Principles, Measurements, and Applications, pages 497-506.Polymers that exhibit higher melt strength have superior processingproperties and can command a higher price.

Polymer characterization is an important branch of chemical analysis.Characterization of a polymer to determine its topology, (in otherwords, the degree and type of branching of the polymer) is currentlyinsufficient for correlating molecular structure to physical properties.Nuclear Magnetic Resonance (NMR) analysis can determine the averagenumber of branch points per polymer molecule; see DePooter, et al., J.App. Pol. Sc., 42, p 399-408 (1991). However, such an NMR analysis doesnot determine the molecular weight distribution of the long chainbranches or the type of branching, for example, “T” branching, “star”branching, “comb” branching and “T” branching.

Polymers have been characterized by Field Flow Fractionation (FFF) byflowing a solution of a polymer in a channel perpendicular to a forcefield (such as a centrifugal force field in a centrifuge) to separatethe components of the polymer in successive elution volumes from thechannel. See, for example, Janca, Field-Flow Fractionation—Analysis ofMacromolecules and Particles, 1988, Marcel Dekker. In FFF, highermolecular weight fractions of the polymer generally elute from thechannel after the lower molecular weight fractions of the polymer. FFFhas not apparently been used to characterize polymers for long chainbranching topology.

Ionic polymers, such as sulfonated polystyrene, have been characterizedby electrophoresis (EP) in a system where the polymer is dissolved in abuffer solution and migrated under the influence of an electric field(electrophoretic mobility) through a medium such as a gel swelled withthe buffer. Lower molecular weight fractions of the polymer migratefaster than higher molecular weight fractions of the polymer. Thecharacterization of long chain branching of ionic polymers has beenattempted using EP but without success. See Smisek and Hoagland,Science, 8 Jun. 1990, p 1221-1223 and especially page 1222, thirdcolumn, which stated: “We next compared the dependence of[electrophoretic] mobility on N [molecular weight] for linear and starPSS [linear and star branched sulfonated polystyrene] (FIG. 4).Surprisingly, over the molecular size range displayed [N from about 100to about 100,000], the mobility depended only on N, and was independentof molecular topology”, in other words, no separation of linear frombranched polymer was observed.

Hydrodynamic Chromatography (HDC) is an important polymercharacterization technique. See, for example, Small, J. ColloidInterface Science, 1974, 48, p 147 and Stegeman et al., J. Chrom., 1993,657(2), p 283-303. In HDC a solution of a polymer is flowed by an eluantover the surfaces of non-porous beads packed in column (or through acapillary column). In HDC the higher molecular weight fractions of thepolymer elute from the column before the lower molecular weightfractions of the polymer. More accurately, HDC separates components of apolymer according to their hydrodynamic size in a solution or adispersion. However, HDC has not apparently been used to characterizepolymers for long chain branching topology.

Size Exclusion Chromatography (SEC) (also called Gel PermeationChromatography (GPC)) is an important polymer characterizationtechnique. See, Yau et al., Modern Size-Exclusion Liquid Chromatography,1979, John Wiley & Sons. In SEC a solution of a polymer is flowed by aneluant through a column packed with porous beads. The polymer diffusesinto and out of the porous beads (there being essentially no flow of theeluant through the porous beads because the flow channels around thebeads are significantly larger than the pores of the beads). In SEC thehigher molecular weight fractions of the polymer elute from the columnbefore the lower molecular weight fractions of the polymer. Moreaccurately, SEC separates components of a polymer according to theirhydrodynamic size.

A branched polymer has a somewhat smaller radius of gyration in solutionthan a linear polymer of the same type and molecular weight. Thus, SECcan be used to characterize a polymer for branching. See Drott andMendelson, Journal of Polymer Science, Part A-2, Vol. 8, 1970, p 1361.However, as pointed out by Drott and Mendelson, as the degree ofbranching of a polymer increases the relative effect on SEC elutionvolume decreases. Furthermore, SEC provides no direct information of theshape of the molecule (for example, star shape or H shape) or themolecular weight of the branch. Thus, the information obtained from SECfor the study of long chain branching of polymers is not sufficient todefine the Theological properties of the polymer. It would be a clearadvance in the art if a better solution were discovered for the problemof characterizing a polymer for long chain branching.

DISCLOSURE OF INVENTION

The instant invention is called “Molecular Topological Fractionation”(MTF). MTF provides a better solution for the problem of characterizinga polymer for long chain branching topology. MTF is a fundamentally newseparation science technique as illustrated by reference to FIG. 1. FIG.1 shows a plot of channel size divided by molecule size on the verticalaxis versus channel flow rate on the horizontal axis. FIG. 1 shows thearea occupied by various polymer characterization techniques, namely:Field Flow Fractionation (FFF), Hydrodynamic Chromatography (HDC), GelPermeation Chromatography (GPC), Electrophoresis (EP) as well as thetechnique of the instant invention (MTF).

As shown in FIG. 1, the channel size divided by the molecule size (themolecule size being expressed as the radius of gyration of the polymerin solution) of GPC and EP can vary from a relatively small value to arelatively large value depending on the pore size and pore size rangeused in the GPC packing or the gel concentration used in EP. However, inboth GPC and EP there is little or no flow of liquid through the poresof the GPC packing or through the gel of the EP system. With regard toFFF, the channel size is large relative to the molecule size and thechannel flow rate is relatively fast. In HDC, the channel size issmaller than in FFF and the channel flow rate is generally (but notnecessarily) relatively fast. In MTF, the channel size is relativelysmall in relation to the molecule size and the channel flow rate isrelatively slow.

More specifically the instant invention is a process for characterizinga sample comprising a population of linear macromolecules of interestand a population of long chain branched macromolecules of interest, theprocess characterized by the following four steps. The first step is toprovide a flow through separating medium and a liquid eluant in whichthe macromolecules of interest dissolve, the separating medium defininga multiplicity of flow through channels which are small enough indiameter so that when the eluant containing dissolved samplemacromolecules of interest is caused to flow under pressure through thechannels of the separating medium, over a certain linear velocity range,a select fraction within the population of the linear macromolecules ofinterest will elute before a select fraction of the population of thelong chain branched macromolecules of interest whereby separate peakelution volumes of said fractions of linear macromolecules and longchain branched macromolecules of interest is established for producingdifferentiation of the fraction of linear macromolecules from thefraction of long chain branched macromolecules, said select fractionsbeing of similar size, the similar size of the fractions of interestbeing characterized so that these fractions will co-elute in a sizeexclusion chromatography experiment optimized so that the fractions areseparated by hydrodynamic size, and wherein the separation is conductedunder temperature and solvent conditions which produce equivalentresults to that obtained when the separation is conducted underisothermal and isocratic conditions. The second step is to introduce asample of dissolved linear macromolecules of interest and long chainbranched macromolecules of interest into the liquid eluant. The thirdstep is to flow the liquid eluant under pressure through the channels ofthe separating medium at a linear velocity that is within the rangespecified in the first step, whereby the sample macromolecules ofinterest emerge from the separating medium separated into successiveelution volumes of the liquid eluant, characterized by said linearmacromolecules of interest eluting before said similarly sized longchain branched macromolecules of interest. The fourth step is todifferentiate the linear macromolecules of interest from the long chainbranched macromolecules of interest based on their successive elutionvolumes established in the third step.

BRIEF DESCRIPTION OF DRAWINGS

FIG. 1 is a plot of channel size divided by molecule size v. channelflow rate for the technique of the instant invention and for variousprior art techniques.

FIG. 2 is a schematic drawing of an apparatus that can be used topractice the process of the instant invention;

FIG. 3 is a cross-sectional microscopic depiction of a porous monolithicpacking porous medium cross-sectioned perpendicular to the direction offlow of a liquid through the porous medium;

FIG. 4 is a cross-sectional microscopic depiction of a particle bedporous medium cross-sectioned perpendicular to the direction of flow ofa liquid through the porous medium;

FIG. 5 is a cross-sectional microscopic depiction of an open cell foamstructure porous medium cross-sectioned perpendicular to the directionof flow of a liquid through the porous medium;

FIG. 6 is a cross-sectional microscopic depiction of a precipitatedsilica type of porous medium cross-sectioned perpendicular to thedirection of flow of a liquid through the porous medium;

FIG. 7 is a plot of molecular weight v. elution volume for linear andlong chain branched polystyrene standards analyzed at a flow rate of tenmicroliters per minute;

FIG. 8 is a plot of molecular weight v. elution volume for linear andlong chain branched polystyrene standards analyzed at a flow rate offifty microliters per minute;

FIG. 9 is a plot of molecular weight v. elution volume for linear andlong chain branched polystyrene standards analyzed at a flow rate of twohundred microliters per minute;

FIG. 10 is a cross-sectional depiction of a chromatographic packingparticle having polymer chains extending from the surface of theparticle; and

FIG. 11 is a schematic depiction summarizing the central features of theprocess of the instant invention.

MODES FOR CARRYING OUT THE INVENTION

Referring now to FIG. 11, the central features of the process of instantinvention will be summarized. As discussed below in greater detail, theinstant invention is a process for characterizing a sample comprising apopulation of linear macromolecules of interest and a population of longchain branched macromolecules of interest. The first step is to providea flow through separating medium 100 and a liquid eluant 101 in whichthe macromolecules of interest dissolve. The separating medium defines amultiplicity of flow through channels 102. A long chain branchedmacromolecule 103 and a linear macromolecule 104 of the same sizedissolved in the eluant 101 are flowed through the channels 102 and thenelute from the separating medium 100 in different elution volumes of theeluant 101 (and if not completely separated, then at least they havedifferent peak elution volumes). The linear macromolecule 104 elutesfirst and is detected as chromatographic peak 105. The long chainbranched macromolecule elutes later and is detected as chromatographicpeak 106.

Referring now to FIG. 2, therein is shown apparatus 10 that can be usedto practice the process of the instant invention. The apparatus 10includes a first reservoir 11 for containing a liquid eluant 12. Theliquid eluant 12 is flowed by first pump 13 through first injectionvalve 14, through a porous separating medium contained in first column15, through first detector 16, through second injection valve 17,through second loop 17 a and then to second waste reservoir 18. Thedetector 16 is responsive to the concentration of polymer in the liquideluant 12, for example, a refractive index detector.

The polymers to be separated (which in general are a blend of linear andlong chain branched polymers, in other words, the polymers to beseparated comprise a population of linear macromolecules of interest anda population of long chain branched macromolecules of interest) aredissolved in a solvent to form a solution of the polymers in a solventand placed in syringe 19 connected to the first injection valve 14 sothat the solution of the polymers in a solvent can be flowed through afirst injection loop 20 and then to a first waste reservoir 21. Thespecific solvent and liquid eluant used in the instant invention is notcritical but preferably the solvent and liquid eluant are the same andare thermodynamically good solvents for the polymers to be separated, inother words, the Mark-Houwink-Sakurada equation alpha exponent valuebeing greater than 0.5 (preferably greater than about 0.7), see Brandrupand Immergut, Polymer Handbook, 1989, John Wiley & Sons. By definitionherein, a long chain branched polymer is a polymer having at least onebranch consisting of at least twenty monomer units (more preferably atleast 100 monomer units and most preferably more than 300 monomerunits). The term “macromolecule” used herein means a synthetic polymersuch as polystyrene or polyethylene; a natural polymer such as apolysaccharide, a polypeptide or a hydrocarbon such as a polyisoprene;or a semisynthetic polymer such as rayon. When the first injection valve14 is rotated from its loop loading position to its loop injectionposition, the solution of the polymers in a solvent contained in thefirst loop 20 is flowed through the porous separating medium containedin the first column 15, through the first detector 16, through thesecond injection valve 17, through second loop 17 a and then to thesecond waste reservoir 18 by the flow of liquid eluant.

Many different types of porous separating media can be used in theinstant invention. For example, the porous separating medium can be aporous monolithic packing, see Gusev et al., J. Chrom. A, 855 (1999)273-290. Porous monolithic packing columns are sold commercially byIsco, Inc. The porous medium can also simply be a bed of particles ofessentially the same size or differing size. The porous medium can alsobe a monolithic packing composed of an open cell foam structure. Theporous medium can also be a porous ceramic material (for example aporous silica or a zeolitic material) having appropriate pore or channelsize.

Although the type of porous separating medium contained in the firstcolumn 15 is not critical in the instant invention, the followingrequirements are necessary. The separating medium must define amultiplicity of flow through channels which are small enough in diameterso that when the eluant containing dissolved sample macromolecules ofinterest is caused to flow under pressure through the channels of theseparating medium, over a certain linear velocity range, a selectfraction within the population of the linear macromolecules of interestwill elute before a select fraction of the population of the long chainbranched macromolecules of interest whereby separate peak elutionvolumes of said fractions of linear macromolecules and long chainbranched macromolecules of interest is established for producingdifferentiation of the fraction of linear macromolecules from thefraction of long chain branched macromolecules, said select fractionsbeing of similar size, the similar size of the fractions of interestbeing characterized so that these fractions will co-elute in a sizeexclusion chromatography experiment optimized so that the fractions areseparated by hydrodynamic size, and wherein the size exclusionchromatography separation is conducted under temperature and solventconditions which produce equivalent results to that obtained when thesize exclusion chromatography separation is conducted under isothermaland isocratic conditions.

The term “isocratic” means that the solvent composition of the eluantused in the size exclusion chromatography separation is constant. Theterm “equivalent” means essentially the same result, in other words,that the fraction of linear macromolecules and the fraction of longchain branched macromolecules of similar size or radius of gyrationco-elute in the size exclusion chromatography experiment. Preferably,the size exclusion chromatography experiment is conducted underisothermal and isocratic conditions.

The linear macromolecules of interest can be differentiated from thelong chain branched macromolecules of interest by quantifying theconcentration of same in each successive elution volume. For example,the refractive index detector 16 is responsive to the concentration ofmacromolecules of interest in the successive elution volumes from thecolumn 15.

Although the diameter of an individual channel of the separating mediumcan be smaller than the radius of gyration of a long chain branchedmacromolecule of interest, at least a portion of the channels need tohave a diameter large enough so that such macromolecule of interest canbe flowed through the channels of the separating medium. On the otherhand, if none of the channels of the separating medium are sufficientlysmall in diameter, then there will not be the required separationbetween the long chain branched macromolecule and the linearmacromolecule. The channels of the separating medium of the instantinvention can have and often will have a range of diameters. However,such range of diameters should not be so large that essentially all ofthe flow of eluant is through large diameter channels that are notsufficiently small to produce the separation between the long chainbranched macromolecule and the linear macromolecule required by theinstant invention.

Preferably, the log of the weight average molecular weight (WAMW) of along chain branched polymer of interest in the sample to becharacterized in grams per mole divided by the effective pore diameter(EPD) in angstroms (log [WAMW÷EPD]) of the porous separating mediumcross-sectioned perpendicular to the direction of flow of the eluantthrough the porous separating medium is in the range of from about 1 toabout 4 and more preferably in the range of from about 2 to about 4.Various methods of determining effective pore diameter are describedbelow.

Described another way, the effective pore diameter of the porousseparating medium cross-sectioned perpendicular to the direction of flowof the eluant through the porous separating medium divided by radius ofgyration of a long chain branched polymer of interest in the sample tobe characterized is preferably in the range of from about 0.5 to about50. As a point of reference, linear polystyrene of ten thousand, onehundred thousand and one million molecular weight has a radius ofgyration in tetrahydrofuran of about three, about thirteen and aboutfifty nanometers respectively, See Yau et al., Modern Size-ExclusionLiquid Chromatography, page 36, 1979, John Wiley & Sons.

Referring now to FIG. 3, therein is shown a cross-sectional electronmicroscopic depiction of a porous monolithic packing porous separatingmedium 50 cross-sectioned perpendicular to the direction of flow ofeluant through the porous medium. The pores 51 in the porous monolithicpacking porous medium 50 are irregularly shaped and of different size.The effective pore diameter of the pores 51 is determined by the averagesize of a circle 52 that can be drawn in the pores 51 excluding anypores that are smaller than 30 Angstroms.

Referring now to FIG. 4, therein is shown a cross-sectional depiction ofa porous separating medium comprising a bed of particles 60cross-sectioned perpendicular to the direction of flow of eluant throughthe porous separating medium. The pores 61 in the bed of particles 60are of uniform size because the particles 60 are of the same size.However, it should be understood that it is not critical in the instantinvention that the particles 60 be essentially the same size. Theeffective pore diameter of the pores 61 is determined by the averagesize of a circle 62 that can be drawn in the pores 61 excluding anypores that are smaller than 30 Angstroms. When the particles 60 are of auniform size, then the effective pore diameter of the pores 61 cansimply be calculated as about 15% of the diameter of the particles 60.

Referring now to FIG. 5, therein is shown a cross-sectional electronmicroscopic depiction of an open cell foam structure 70 porousseparating medium cross-sectioned perpendicular to the direction of flowof eluant through the porous separating medium. The effective porediameter of the pores 71 in the structure 70 is determined by theaverage size of a circle 72 that can be drawn in the pores 71 excludingany pores that are smaller than 30 Angstroms. The pores 71 are shown asbeing circular in FIG. 4 for simplicity. However, in reality the pores71 are often polygonal in shape.

Referring now to FIG. 6, therein is shown a cross-sectional schematicdepiction of a silica porous separating medium cross-sectionedperpendicular to the direction of flow of eluant through the porousseparating medium. The silica porous separating medium comprises silicaspheres 80 connected by silica strands 81. The effective pore diameterof the pores in the silica porous separating medium is determined by theaverage size of a circle 82 that can be drawn in the pores. The silicaporous separating medium of the type shown in FIG. 6 can be made by theprocess described in U.S. Pat. No. 4,112,032 including hydrothermaltreatment to increase mechanical strength and to improve pore sizedistribution as described by Iler, The Chemistry of Silica, Wiley, 1979,p 539-544.

Referring now to FIG. 10, therein is shown a cross-sectional depictionof a chromatographic packing particle 90 useful in the instantinvention, the packing particle 90 having polymer chains 91 extendingfrom the surface of the particle. Each of the polymer chains 91 mostpreferably consist of a minimum of 100 monomer units. Preferably, morethan one half of the polymer chains 91 consist of a minimum of 300monomer units. The polymer chains 91 can be attached to the particle 90by the procedure outlined by von Verne and Patten, J. Am. Chem. Soc.1999, 121, 7409-7410. The particle 90 can be any convenient particlesuch as silica or a synthetic polymer. It should also be understood thatthe exposed surfaces of a monolithic packing can also be treated so thatsuch polymer chains extend from the surfaces of such a monolithicpacking.

Referring still to FIG. 10, most preferably the kind of polymer of thepolymer chains 91 is the same as the long chain branched polymer to becharacterized. For example, if the long chain branched polymer to becharacterized is polystyrene, then the polymer chains 91 arepolystyrene. Preferably, the polymer of the polymer chains 91 ismiscible with the solution of long chain branched polymer to becharacterized.

Referring again to FIG. 2, as the solution of the polymers is flowedthrough the porous separating medium contained in the column 15 by theflow of the liquid eluant 12 therethrough, the elution volume of thepolymer will be a function of the molecular weight of the polymer, theflow rate of the liquid eluant 12 and the topology of the polymer.Referring now to FIG. 7, therein is shown a plot of log molecular weight(in terms of linear polystyrene equivalent weight average molecularweight in gram moles) v. elution volume for linear and long chainbranched polystyrene samples analyzed at a flow rate of the liquid 12(of FIG. 2) of ten microliters per minute (a linear velocity of 0.6millimeters per minute) using a 4.6 millimeter internal diameter byfifty millimeter long column 15 (of FIG. 2) containing porous monolithicpacking porous separating medium having an effective pore diameter ofabout 1,000 Angstroms. The square data points represent the peak elutionvolumes for the linear polystyrene samples. The dot data pointsrepresent the peak elution volumes of the long chain branchedpolystyrene samples.

It should be understood that the term “linear velocity” used hereinmeans the linear velocity of the eluant in the chromatography columnassuming that the column is not packed and assuming plug flow. Referringnow to FIG. 2, of course, the actual linear velocity of the liquid 12through the monolithic porous separating medium in column 15 is fasterbecause the fraction of space in the column that comprises flow channelsis less than the volume of the column 15. Never-the-less, the abovestated definition of “linear velocity” is used herein for simplicity andto be consistent with the use of this term in the literature of LiquidChromatography as the “superficial or empty-tube velocity”, see J.Chromatography Library, Vol. 22A, page A38.

Referring now to FIG. 8, therein is shown a plot of log molecular weight(in terms of linear polystyrene equivalent weight average molecularweight in gram moles) v. elution volume for the linear and long chainbranched polystyrene samples analyzed at a flow rate of the liquid 12(of FIG. 2) of fifty microliters per minute (a linear velocity of 3millimeters per minute) using the same 4.6 millimeter internal diameterby fifty millimeter long column 15 containing porous monolithic packingporous separating medium as FIG. 7. The square data points representpeak elution volumes for the linear polystyrene samples. The dot datapoints represent peak elution volumes for the long chain branchedpolystyrene samples.

Referring now to FIG. 9, therein is shown a plot of log molecular weight(in terms of linear polystyrene equivalent weight average molecularweight in gram moles) v. elution volume for linear and long chainbranched polystyrene standards analyzed at a flow rate of the liquid 12(of FIG. 2) of two hundred microliters per minute (a linear velocity of12 millimeters per minute) using the same 4.6 millimeter internaldiameter by fifty millimeter long column as FIGS. 7 and 8. The squaredata points represent peak elution volumes for the linear polystyrenesamples. The dot data points represent the peak elution volumes for thelong chain branched polystyrene samples.

Referring still to FIG. 9, all of the data points lay on a straight linejust as expected as if the mechanism of separation in the column 15 (ofFIG. 2) were Hydrodynamic Chromatography. Referring now to FIG. 7, itwill be noted that for log weight average molecular weight above about5, there are no data points for long chain branched polymer. Long chainbranched polymer having a log weight average molecular weight aboveabout 5 may have become trapped in the column 15 (of FIG. 2) under theseconditions (or at least such polymer has an elution volume greater than0.45 milliliters).

When using the relatively fast flow rate of FIG. 9, there is no apparentseparation of the linear and the long chain branched polymers on thecolumn 15 (of FIG. 2). When using the intermediate flow rate of FIG. 8,there is a substantial and surprising separation between the linear andthe long chain branched polymers. The optimum flow rate of the liquideluant 12 (of FIG. 2) in the instant invention in order to obtain theoptimum separation of polymers depends, of course, on the specificsystem used and can be relatively easily determined by experimentation.

It is believed that the instant invention can be used to separate notonly linear (and short chain branched) polymer from long chain branchedpolymer but also two different long chain branched polymers even if theyhave the same molecular weight and are the same type provided they havedifferent topology. For example, it is believed that the instantinvention can be used to separate a “T” topology polystyrene of onemillion molecular weight from an “H” topology polystyrene of one millionmolecular weight.

Referring to FIG. 8, the porous separating medium used in the column 15(of FIG. 2) to generate the data shown does not appear to be effectivefor log weight average molecular weights less than about 5. However, ifthe effective pore diameter of the porous medium used in the column 15(of FIG. 2) is reduced, then the process of the instant invention can beused to characterize samples having long, chain branched polymer ofinterest of lower log weight average molecular weight.

Referring still to FIG. 8, polymer eluting at an elution volume ofgreater than about 0.34 milliliters may be an indication of a relativelyhigh molecular weight long chain branched polymer or a relatively lowermolecular weight polymer. This dichotomy is preferably solved bysubjecting the successive volumes of the liquid eluant 12 (of FIG. 2)flowing from the detector 16 (of FIG. 2) to a separation techniquecapable of separating macromolecules based on molecular size, forexample, size exclusion chromatography (SEC).

Referring now to FIG. 2, SEC eluant 22, for example, tetrahydrofuran,contained in SEC eluant reservoir 23 is pumped by second pump 24 throughsecond injection valve 17, through SEC column 25, through secondrefractive index detector 26, through third detector 27 (such as alow-angle light scattering detector or a viscosity measuring detector)and then to third waste reservoir 28. First detector 16, second detector26 and third detector 27 are connected to a general-purpose digitalcomputer 29 by way of wires 30 (wireless communication is, of course,also suitable). The second injection valve 17 is periodically used toinject a successive elution volume of the liquid 12 onto the sizeexclusion chromatography column 25 to determine the hydrodynamic volumedistribution of a polymer dissolved in the successive elution volume ofthe liquid 12. The relatively low flow rate of the liquid 12 through thecolumn 15 permits a plurality of such SEC determinations during a singleelution of a mixture of polymers from the column 15.

Referring still to FIG. 2, the computer 29 is programmed to periodicallyactuate the second injection valve 17 and to interpret and deconvolutethe signals from the detectors 16, 26 and 27. For example, referring nowto FIG. 7, polymer eluting at 0.40 milliliters can be determined to be6.2 log weight average molecular weight long chain branched polymerand/or 4.0 log weight average molecular weight linear polymer becausethese two polymers have substantially different hydrodynamic volume asmeasured by SEC. When SEC is coupled with MTF, then it is preferablethat the MTF system precede the SEC system as shown in FIG. 2. However,it should be understood that the SEC system can also precede the MTFsystem.

Scanning probe microscopy is a powerful technique for thecharacterization of the shape or topology of a single macromolecule, seeGerle et al., Macromolecules, 1999, 32, 2629-2637 and Prokhorova et al.,Macromolecules, 1999, 32, 2653-2660. Referring to FIG. 2, themacromolecules of interest eluting from the size exclusionchromatography column 25 can be characterized by scanning probemicroscopy in the instant invention by collecting a fraction of theeluant from the column 25 and then subjecting such fraction to scanningprobe microscopy.

The process of the instant invention can be used to develop or produce apolymer, the polymer comprising long chain branched polymer. Forexample, different polymerization conditions (such as temperature,monomer feed rates, pressure, catalyst type and amount) can be studiedto determine which conditions will produce a desired amount and type oflong chain branching and the full scope of the instant inventionincludes the polymer so developed or produced. Similarly, the process ofthe instant invention can be used to develop a polymerization catalyst,the polymerization catalyst optimized to catalyze the polymerization ofa polymer comprising long chain branched polymer. The full scope of theinstant invention includes the catalyst so developed. For example,polymer produced by metalocene polymerization catalysts produced bycombinatorial techniques can be analyzed using the process of theinstant invention to identify the catalysts that produce the desiredamount and type of long chain branching.

1. A process for characterizing a sample comprising a population oflinear macromolecules of interest and a population of long chainbranched macromolecules of interest, the process characterized by thesteps of: (a) providing a sample comprising a population of linearmacromolecules of interest comprising a select fraction (104) and apopulation of long chain branched macromolecules of interest comprisinga select fraction (103), said select fractions being of similar size,the similar size of the fractions of interest being characterized sothat these fractions will co-elute in a size exclusion chromatographyexperiment optimized so that the fractions are separated by hydrodynamicsize and the separation is conducted under temperature and solventconditions which produce equivalent results to that obtained when theseparation is conducted under isothermal and isocratic conditions; (b)providing a flow through separating medium (100) and a liquid eluant(101) in which the macromolecules of interest dissolve, the separatingmedium defining a multiplicity of flow through channels (102), which aresmall enough in diameter so that when the eluant containing dissolvedsample macromolecules of interest is caused to flow under pressurethrough the channels (102) over a certain linear velocity range, theselect fraction (104) of the population of the linear macromolecules ofinterest will elute before the select fraction (103) of the populationof the long chain branched macromolecules of interest whereby separatepeak elution volumes (105) and (106) of said fractions of linearmacromolecules and long chain branched macromolecules of interest areestablished for producing differentiation of the fraction of linearmacromolecules from the fraction of long chain branched macromolecules;(c) determining said linear velocity range; (d) introducing the sampleof dissolved linear macromolecules of interest and long chain branchedmacromolecules of interest into the liquid eluant; (e) flowing theliquid eluant under pressure through the channels of the separatingmedium at a linear velocity that is within the range specified in step(c), whereby the sample macromolecules of interest emerge from theseparating medium separated into successive elution volumes of theliquid eluant, characterized by said linear macromolecules of interesteluting (105) before said similarly sized long chain branchedmacromolecules of interest (106); and (f) differentiating the linearmacromolecules of interest from the long chain branched macromoleculesof interest based on their successive elution volumes established instep (e).
 2. The process of claim 1 wherein the linear macromolecules ofinterest are differentiated from the long chain branched macromoleculesof interest by quantifying the concentration of same in each successiveelution volume.
 3. The process of claim 1 wherein said linear velocityrange is determined on the lower end of the range by a velocity at whichthe sample macromolecules fail to elute from the separating medium, andat the upper end of the range by a velocity at which the long chainbranched macromolecules elute in the same elution volume as the linearmacromolecules of interest so that said linear macromolecules ofinterest cannot be differentiated from said long branched macromoleculesof interest based on peak elution volume.
 4. The process of claim 3wherein the ratio of the peak elution volume of the long chain branchedmacromolecules of interest in the eluant to the peak elution volume ofthe dissolved linear macromolecules of interest in the eluant is greaterthan
 1. 5. The process of claim 1 wherein a successive elution volumefrom step (e) is subjected to size exclusion chromatography to separateany macromolecules present based on hydrodynamic size.
 6. The process ofclaim 5 wherein the macromolecules of interest eluting from the sizeexclusion chromatography separation medium are detected based on theirconcentration.
 7. The process of claim 5 wherein the macromolecules ofinterest eluting from the size exclusion chromatography separationmedium are detected by light scattering.
 8. The process of claim 5wherein the macromolecules of interest eluting from the size exclusionchromatography separation medium are detected by viscometry.
 9. Theprocess of claim 5 wherein the macromolecules of interest eluting fromthe size exclusion chromatography separation medium are detected basedon multi-angle light scattering.
 10. The process of claim 5 wherein theshape of the macromolecules of interest eluting from the size exclusionchromatography separating medium is characterized by scanning probemicroscopy.